The natural furanone (5Z)-4-bromo-5-(bromomethylene)-3-butyl-2(5H)-furanone disrupts quorum sensing-regulated gene expression in Vibrio harveyi by decreasing the DNA-binding activity of the transcriptional regulator protein luxR

AbstractThe tumor suppressor p53 protein activates expression of a vast gene network in response to stress stimuli for cellular integrity. The molecular mechanism underlying how p53 targets RNA polymerase II (Pol II) to regulate transcription remains unclear. To elucidate the p53/Pol II interaction, we have determined a 4.6 Å resolution structure of the human p53/Pol II assembly via single particle cryo-electron microscopy. Our structure reveals that p53’s DNA binding domain targets the upstream DNA binding site within Pol II. This association introduces conformational changes of the Pol II clamp into a further-closed state. A cavity was identified between p53 and Pol II that could possibly host DNA. The transactivation domain of p53 binds the surface of Pol II’s jaw that contacts downstream DNA. These findings suggest that p53’s functional domains directly regulate DNA binding activity of Pol II to mediate transcription, thereby providing insights into p53-regulated gene expression.

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Extinction of insulin gene expression in hybrids between beta cells and fibroblasts is accompanied by loss of the putative beta-cell-specific transcription factor IEF1

Molecular and Cellular Biology ◽

10.1128/mcb.11.3.1547-1552.1991 ◽

1991 ◽

Vol 11 (3) ◽

pp. 1547-1552

Author(s):

D Leshkowitz ◽

M D Walker

Keyword(s):

Gene Expression ◽

Dna Binding ◽

Binding Activity ◽

Insulin Gene ◽

Hybrid Cells ◽

Dna Binding Activity ◽

Insulin Producing Cells ◽

Insulin Gene Expression ◽

Insulinoma Cells

Insulin-producing cells and fibroblasts were fused to produce hybrid lines. In hybrids derived from both hamster and rat insulinoma cells, no insulin mRNA could be detected in any of seven lines examined by Northern (RNA) analysis despite the presence in each line of the insulin genes of both parental cells. Hybrid cells were transfected with recombinant chloramphenicol acetyltransferase plasmids containing defined segments of the rat insulin I gene 5' flank. We observed no transcriptional activity of the intact insulin enhancer or of IEB2, a critical cis-acting element of the insulin enhancer. IEB2 has previously been shown to interact in vitro with IEF1, a DNA-binding activity observed selectively in insulin-producing cells. Hybrid cells showed no detectable IEF1 activity. Furthermore, the insulin enhancer was unable to reduce transcription directed by the Moloney sarcoma virus enhancer in a double-enhancer construct. Thus, extinction of insulin gene expression in the hybrids apparently does not operate through a direct action of repressors on the insulin enhancer; rather, extinction is accompanied by, and may be caused by, reduced DNA-binding activity of the putative transcriptional activator IEF1.

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Cinnamaldehyde and cinnamaldehyde derivatives reduce virulence in Vibrio spp. by decreasing the DNA-binding activity of the quorum sensing response regulator LuxR

BMC Microbiology ◽

10.1186/1471-2180-8-149 ◽

2008 ◽

Vol 8 (1) ◽

pp. 149 ◽

Cited By ~ 173

Author(s):

Gilles Brackman ◽

Tom Defoirdt ◽

Carol Miyamoto ◽

Peter Bossier ◽

Serge Van Calenbergh ◽

...

Keyword(s):

Dna Binding ◽

Quorum Sensing ◽

Response Regulator ◽

Binding Activity ◽

Dna Binding Activity ◽

Vibrio Spp

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Overexpression of mitogen-activated protein kinase (ERK1) enhances T- cell cytokine gene expression: role of AP1, NF-AT, and NF-KB

Blood ◽

10.1182/blood.v82.8.2470.2470 ◽

1993 ◽

Vol 82 (8) ◽

pp. 2470-2477 ◽

Cited By ~ 36

Author(s):

JH Park ◽

L Levitt

Keyword(s):

Gene Expression ◽

T Cell ◽

Dna Binding ◽

Map Kinase ◽

Binding Activity ◽

Cytokine Gene Expression ◽

Cytokine Gene ◽

Dna Binding Activity ◽

Mitogen Activated Protein

Abstract Transfected Jurkat cells overexpressing extracellular signal-regulated kinase (ERK1), also referred to as mitogen-activated protein (MAP) kinase, were selected by Western blotting assay using anti-ERK1 and antiphosphotyrosine antibodies in combination with a functional MAP kinase assay. We then asked whether enhanced ERK1 expression had any effect on induction of T-cell cytokine genes. The results show that overexpression of ERK1 enhances expression of T-cell interleukin-2 (IL- 2), IL-3, and granulocyte-macrophage colony-stimulating factor mRNA; no change was seen in expression of the alpha-actin gene. DNA-binding activities of the transcription factors AP1, NF-AT, and NF-kB were specifically increased twofold to fourfold in ERK1-overexpressing clones relative to nontransformed or vector-transformed cells, whereas no enhancement of CK1-CK2 protein DNA binding activity was detected after ERK1 overexpression. Additionally, increased NF-AT DNA binding activity was associated with functional enhancement of NF-AT transactivating activity in ERK1-overexpressing cells. These results provide direct evidence for the role of MAP kinase in the regulation of cytokine gene expression and indicate that such regulation is likely mediated through the enhanced DNA binding activity of specific nuclear transcription factors.

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Abstract P2-09-10: Antiprogestin CDB4124 suppressed progesterone receptor-mediated DNA binding activity, proliferation and gene expression in T47D cancer cells and normal human breast microstructures

10.1158/0008-5472.sabcs13-p2-09-10 ◽

2013 ◽

Author(s):

A Gupta ◽

D Ivancic ◽

J Wang ◽

E Horvath ◽

O Lee ◽

...

Keyword(s):

Gene Expression ◽

Dna Binding ◽

Progesterone Receptor ◽

Cancer Cells ◽

Binding Activity ◽

Human Breast ◽

Dna Binding Activity ◽

Normal Human Breast ◽

Normal Human

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Analysis of Activator and Repressor Functions Reveals the Requirements for Transcriptional Control by LuxR, the Master Regulator of Quorum Sensing in Vibrio harveyi

mBio ◽

10.1128/mbio.00378-13 ◽

2013 ◽

Vol 4 (4) ◽

Cited By ~ 43

Author(s):

Julia C. van Kessel ◽

Luke E. Ulrich ◽

Igor B. Zhulin ◽

Bonnie L. Bassler

Keyword(s):

Gene Expression ◽

Transcription Factors ◽

Dna Binding ◽

Quorum Sensing ◽

Binding Site ◽

Binding Sites ◽

Vibrio Harveyi ◽

Communication Process ◽

Nucleotide Sequencing ◽

Collective Behaviors

ABSTRACT LuxR-type transcription factors are the master regulators of quorum sensing in vibrios. LuxR proteins are unique members of the TetR superfamily of transcription factors because they activate and repress large regulons of genes. Here, we used chromatin immunoprecipitation and nucleotide sequencing (ChIP-seq) to identify LuxR binding sites in the Vibrio harveyi genome. Bioinformatics analyses showed that the LuxR consensus binding site at repressed promoters is a symmetric palindrome, whereas at activated promoters it is asymmetric and contains only half of the palindrome. Using a genetic screen, we isolated LuxR mutants that separated activation and repression functions at representative promoters. These LuxR mutants exhibit sequence-specific DNA binding defects that restrict activation or repression activity to subsets of target promoters. Altering the LuxR DNA binding site sequence to one more closely resembling the ideal LuxR consensus motif can restore in vivo function to a LuxR mutant. This study provides a mechanistic understanding of how a single protein can recognize a variety of binding sites to differentially regulate gene expression. IMPORTANCE Bacteria use the cell-cell communication process called quorum sensing to regulate collective behaviors. In vibrios, LuxR-type transcription factors control the quorum-sensing gene expression cascade. LuxR-type proteins are structural homologs of TetR-type transcription factors. LuxR proteins were assumed to function analogously to TetR proteins, which typically bind to a single conserved binding site to repress transcription of one or two genes. We find here that unlike TetR proteins, LuxR acts a global regulator, directly binding upstream of and controlling more than 100 genes. Again unlike TetR, LuxR functions as both an activator and a repressor, and these two activities can be separated by mutagenesis. Finally, the consensus binding motifs driving LuxR-activated and -repressed genes are distinct. This work shows that LuxR, although structurally similar to TetR, has evolved unique features enabling it to differentially control a large regulon of genes in response to quorum-sensing cues.

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Global H-NS counter-silencing by LuxR activates quorum sensing gene expression

Nucleic Acids Research ◽

10.1093/nar/gkz1089 ◽

2019 ◽

Cited By ~ 6

Author(s):

Ryan R Chaparian ◽

Minh L N Tran ◽

Laura C Miller Conrad ◽

Douglas B Rusch ◽

Julia C van Kessel

Keyword(s):

Gene Expression ◽

Quorum Sensing ◽

Cell Communication ◽

Binding Activity ◽

Primary Role ◽

Dna Binding Activity ◽

Cellular Behaviors ◽

A Cell ◽

Promoter Dna

Abstract Bacteria coordinate cellular behaviors using a cell–cell communication system termed quorum sensing. In Vibrio harveyi, the master quorum sensing transcription factor LuxR directly regulates >100 genes in response to changes in population density. Here, we show that LuxR derepresses quorum sensing loci by competing with H-NS, a global transcriptional repressor that oligomerizes on DNA to form filaments and bridges. We first identified H-NS as a repressor of bioluminescence gene expression, for which LuxR is a required activator. In an hns deletion strain, LuxR is no longer necessary for transcription activation of the bioluminescence genes, suggesting that the primary role of LuxR is to displace H-NS to derepress gene expression. Using RNA-seq and ChIP-seq, we determined that H-NS and LuxR co-regulate and co-occupy 28 promoters driving expression of 63 genes across the genome. ChIP-PCR assays show that as autoinducer concentration increases, LuxR protein accumulates at co-occupied promoters while H-NS protein disperses. LuxR is sufficient to evict H-NS from promoter DNA in vitro, which is dependent on LuxR DNA binding activity. From these findings, we propose a model in which LuxR serves as a counter-silencer at H-NS-repressed quorum sensing loci by disrupting H-NS nucleoprotein complexes that block transcription.

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Signaling mechanisms regulating bombesin-mediated AP-1 gene induction in the human gastric cancer SIIA

AJP Cell Physiology ◽

10.1152/ajpcell.2000.279.2.c326 ◽

2000 ◽

Vol 279 (2) ◽

pp. C326-C334 ◽

Cited By ~ 41

Author(s):

Hong Jin Kim ◽

B. Mark Evers ◽

David A. Litvak ◽

Mark R. Hellmich ◽

Courtney M. Townsend

Keyword(s):

Gene Expression ◽

Gastric Cancer ◽

Protein Kinase ◽

Dna Binding ◽

Signaling Pathways ◽

Binding Activity ◽

Human Gastric Cancer ◽

Rapid Induction ◽

Dna Binding Activity ◽

Jun B

The hormone bombesin (BBS) and its mammalian equivalent gastrin-releasing peptide (GRP) act through specific GRP receptors (GRP-R) to affect multiple cellular functions in the gastrointestinal tract; the intracellular signaling pathways leading to these effects are not clearly defined. Previously, we demonstrated that the human gastric cancer SIIA possesses GRP-R and that BBS stimulates activator protein-1 (AP-1) gene expression. The purpose of our present study was to determine the signaling pathways leading to AP-1 induction in SIIA cells. A rapid induction of c- jun and jun-B gene expression was noted after BBS treatment; this effect was blocked by specific GRP-R antagonists, indicating that BBS is acting through the GRP-R. The signaling pathways leading to increased AP-1 gene expression were delineated using phorbol 12-myristate 13-acetate (PMA), which stimulates protein kinase C (PKC)-dependent pathways, by forskolin (FSK), which stimulates protein kinase A (PKA)-dependent pathways, and by the use of various protein kinase inhibitors. Treatment with PMA stimulated AP-1 gene expression and DNA binding activity similar to the effects noted with BBS; FSK stimulated jun-B expression but produced only minimal increases of c- jun mRNA and AP-1 binding activity. Pretreatment of SIIA cells with either H-7 or H-8 (primarily PKC inhibitors) inhibited the induction of c- jun and jun-B mRNAs in response to BBS, whereas H-89 (PKA inhibitor) exhibited only minimal effects. Pretreatment with tyrphostin-25, a protein tyrosine kinase (PTK) inhibitor, attenuated the BBS-mediated induction of c- jun and jun-B, but the effect was not as pronounced as with H-7. Collectively, our results demonstrate that BBS acts through its receptor to produce a rapid induction of both c- jun and jun-B mRNA and AP-1 DNA binding activity in the SIIA human gastric cancer. Moreover, this induction of AP-1, in response to BBS, is mediated through both PKC- and PTK-dependent signal transduction pathways with only minimal involvement of PKA.

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Efficient transcription of the glycolytic gene ADH1 and three translational component genes requires the GCR1 product, which can act through TUF/GRF/RAP binding sites

Molecular and Cellular Biology ◽

10.1128/mcb.10.2.859-862.1990 ◽

1990 ◽

Vol 10 (2) ◽

pp. 859-862

Author(s):

G M Santangelo ◽

J Tornow

Keyword(s):

Gene Expression ◽

Saccharomyces Cerevisiae ◽

Dna Binding ◽

Binding Sites ◽

Binding Activity ◽

Heterologous Gene ◽

Dna Binding Activity ◽

Glycolytic Gene

Glycolytic gene expression in Saccharomyces cerevisiae is thought to be activated by the GCR and TUF proteins. We tested the hypothesis that GCR function is mediated by TUF/GRF/RAP binding sites (UASRPG elements). We found that UASRPG-dependent activation of a heterologous gene and transcription of ADH1, TEF1, TEF2, and RP59 were sensitive to GCR1 disruption. GCR is not required for TUF/GRF/RAP expression or in vitro DNA-binding activity.

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NF-kB as a Regulator of FA/BRCA Gene Expression in Multiple Myeloma.

Blood ◽

10.1182/blood.v110.11.3508.3508 ◽

2007 ◽

Vol 110 (11) ◽

pp. 3508-3508

Author(s):

Vasco A. Oliveira ◽

Linda Mathews ◽

Danielle Yarde ◽

Xingyu Wang ◽

David Boulware ◽

...

Keyword(s):

Gene Expression ◽

Multiple Myeloma ◽

Drug Resistance ◽

Dna Binding ◽

Cell Lines ◽

Myeloma Cell ◽

Binding Activity ◽

Acquired Drug Resistance ◽

Brca Gene ◽

Dna Binding Activity

Abstract Results to date argue compellingly that disruption of FA/BRCA gene expression plays a pivotal role in human somatic carcinogenesis. Melphalan, a DNA cross-linker, is one of the most widely used and effective drugs in the treatment of multiple myeloma (MM). Although most patients respond to standard and high dose melphalan, eventually patients acquire resistance and develop progressive disease. In 1991, our laboratory reported that acquired resistance in a human myeloma cell line was associated with reduced DNA crosslinks, elevated glutathione levels, and increased radiation survival (Cancer Res. 5:993; 1991). Most recently, we reported that the melphalan-resistant myeloma cell lines, 8226/LR5 and U266/LR6, showed a significant increase in several FA/BRCA genes compared to drug-sensitive cells, and that enhanced interstrand crosslink (ICL) repair via this signaling pathway contributes to acquired drug resistance in melphalan resistant cell lines (Blood 10:698; 2005). Here, we report that IKKa is constitutively phosphorylated in unstimulated 8226/LR5 cells, but not in melphalan-sensitive control cells. The specific phosphorylation of IKKa leads to an increase in basal NF-kB DNA binding activity, and 8226/LR5 cells are found to be markedly sensitive to BMS-345541 (a highly selective inhibitor of IkB) relative to control cells. Importantly, a cytotoxic dose of BMS-345541 induces a dramatic decrease in FA/BRCA gene expression, and a concomitant inhibition of NF-kB DNA binding activity in both 8226/S and 8226/LR5 cells. Furthermore, we show that 8226/LR5 cells experience the highest degree of direct binding between FANCD2 promoter and NF-kB/Rel family members, which, in turn, leads to an increase in basal FANCD2-specific NF-kB activity. Small-interfering RNA (siRNA)-mediated depletion of RelB and p50, but not other NF-kB subunits, in 8226 cells results in impaired NF-kB binding activity, and visible decrease in FANCD2 protein expression. Studies designed to dissect the role of NF-kB in acquired melphalan resistance are in progress, and the results will be presented. Our findings suggest that NF-kB functions as a regulator of FA/BRCA expression, and that this pathway represents a new target for preventing acquired drug resistance in myeloma patients.

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